Site‐specific Recombination: Uses in Biotechnology

Abstract

Site‐specific recombination has become an important tool for manipulating DNA both in vitro and in vivo. It has found widespread use in controlling gene expression in transgenic plants and animals.

Keywords: recombination; site‐specific; phage display; transgenics; cre‐lox

Figure 1.

Use of the Cre‐lox site‐specific recombination system in bacteriophage λ cDNA cloning vectors. The vector shown at the top has a high copy plasmid inserted between loxP sites bounded by the left and right arms of the λ genome. The cDNA is cloned into the plasmid and the DNA packaged in λ particles. These phages can be used to infect cells expressing the Cre recombinase which promotes the recombination of the loxP sites. The plasmid is excised and free to replicate autonomously.

Figure 2.

Increasing molecular diversity using site‐specific recombination. (a) Phage fd containing a library of VH genes fused to the minor coat protein gIII are used to infect cells with plasmids containing a library of VL chains. (b) In the presence of Cre recombinase, the phage and plasmid undergo recombination via the loxP or loxP511 sites to yield the two possible cointegrate structures shown. If the identical lox sites used to form the cointegrates recombine a second time the starting structures shown in (a) will result. (c) If the second set of lox sites in the cointegrate structures are recombined, the structure pairing both libraries will be formed.

Figure 3.

Generation of conditional knockout mice using site‐specific recombination. (a) Targeted construct, gene A, flanked by loxP sites and a selectable marker are introduced by homologous recombination into the genome of ES cells. Once integrated, Cre is expressed transiently in these cells to remove the selectable marker via lox recombination. (b) ES cells containing gene A flanked by loxP sites are introduced into mice. The resulting mice are crossed with mice containing a cre gene whose expression can be controlled conditionally. (c) Offspring will then excise gene A when Cre is expressed.

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References

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Further Reading

Hoess RH and Abremski K (1990) The Cre‐lox recombination system. In: Eckstein F and Lilley DMT (eds) Nucleic Acids and Molecular Biology, vol. 4, pp. 99–109. Berlin: Springer‐Verlag.

Jiang R and Gridley T (1997) Gene targeting: Things go better with Cre. Current Biology 7: R321–R323.

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How to Cite close
Hoess, Ronald H(Apr 2001) Site‐specific Recombination: Uses in Biotechnology. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1038/npg.els.0001060]